A key factor in the recent advances in molecular biology has been the development of reliable and convenient methods for synthesizing polynucleotides, e.g. Itakura, Science, Vol. 209, pgs. 1401-1405 (1980); and Wallace et al, pgs. 631-663, in Scouten, ed. Solid Phase Biochemistry (John Wiley & Sons, New York, 1982). As the use of synthetic polynucleotides has increased, the demand for even greater convenience in the preparation of pure, ready-to-use polynucleotides has also increased. This demand has stimulated the development of many improvements in the basic procedures for solid phase synthesis, e.g. Sinha et al, Nucleic Acids Research, Vol. 12, pgs. 4539-4557 (1984)(beta-cyanoethyl in phosphoramidite chemistries); Froehler et al, Tetrahedron Letters, Vol. 27, pgs. 469-472 (1986)(H-phosphonate chemistry); Germann et al, Anal. Biochem., Vol. 165, pgs. 399-405 (1987); and Ikuta et al, Anal. Chem., Vol. 56, pgs. 2253-2256 (1984)(rapid purification of synthetic oligonucleotides by way of trityl moieties); Molko et al, European patent publication 241363 dated 3 April 1987 (improved base-labile acyl protection groups for exocyclic amines), and the like.
In spite of such progress, difficulties are still encountered in current methods of polynucleotide synthesis and purification. For example, derivatized controlled pore glass (CPG), the current support of choice in most solid phase methodologies, can be responsible for spurious indications of coupling yields, e.g. Pon et al, BioTechniques, Vol. 6, pgs. 768-775 (1988). Moreover, CPG, like most glasses, lacks chemical stability in some of the highly corrosive deprotection reagents, such as concentrated ammonia and trichloroacetic acid, used in polynucleotide synthesis. As a consequence, the CPG support itself can be degraded in the deprotection steps and can be a source of contamination of the final product. This problem is exacerbated by the relative long reaction times required to remove currently used protection groups for exocyclic amines. An extended period of deprotection is required to remove these groups after the polynucleotide has been cleaved from the solid phase support. Thus, complete automation of synthesis and purification has been impractical. Another problem with CPG is that its surface supports chain growth at sites other than those associated with the 5' terminus of an attached nucleoside. Such "extraneous" chain growth gives rise to a heterogeneous population of 5'-blocked (usually tritylated) polynucleotides. Typically, the "extraneous" tritylated products lack one or more 3' nucleotides. This, of course, prevents one from successfully taking advantage of the relatively high hydrophobicity of the trityl group to purify "correct sequence" polynucleotides. Incorrect-sequence extraneous chains are also tritylated.
In view of the above, the field of solid phase polynucleotide synthesis could be significantly advanced by the availability of alternative support materials which have the favorable mechanical properties of CPG, but which also possess greater chemical stability under the reaction conditions of polynucleotide synthesis, and which provide less opportunity for extraneous chain growth during synthesis. The use of such materials coupled with improved exocyclic protection groups would allow practical automation of polynucleotide synthesis and purification in a single instrument.